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Results for “cell biology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 343 records · Page 19Linked to original sources

bioRxiv2025

Deep-learning deconvolution and segmentation of fluorescent membranes for high-precision bacterial cell-size profiling

Evolutionary studies in bacteria have emphasized genetic and metabolic diversity, while cell-size variation has received less attention. Here we introduce MEDUSSA, a high-throughput method for precise bacterial cell-size profiling based on automatic segmentation of fluorescent membrane images, well suited to studying cell-size diversity. The approach uses deep-learning-based membrane deconvolution, segmentation models fine-tuned for fluorescent-membrane images, and error-corrected cell measurement to extract accurate sizes from individual bacterial cells regardless of shape, size, chaining, or clustering. Our method overcomes limitations of phase-contrast segmentation, yielding reliable single-cell dimensions. We applied MEDUSSA to six strains of Priestia megaterium and found over twofold differences in cell volume across strains, largely driven by differences in cell width. We identified a partially-functional PBP1 allele that underlies the reduced width of one strain. Together, these results demonstrate the power of comparative analyses in bacterial cell biology and expand the toolkit to investigate the evolution of bacterial cell size.

cell biology↗

bioRxiv2026

A gene-agnostic FACS technique to isolate stem cells in Hydractinia symbiolongicarpus validated with cytology

Hydractinia symbiolongicarpus is a powerful model for stem cell research and maintains a population of pluripotent adult stem cells throughout its lifetime. Here we describe a gene expression-agnostic FACS technique to isolate a live cell population from Hydractinia feeding polyps that appear to be stem cells. This technique utilizes only the general cellular component stains DAPI, DRAQ5, Calcein AM, and Pyronin Y. The stem cell population was identified via subtractive gating based on samples whose stem cell populations had been selectively depleted with the DNA-alkylating agent Mitomycin C. To validate the identity of the isolated population, a colorimetric cytological assay capable of simultaneously discriminating between all major Hydractinia cell types in a live-dissociated cell solution was developed using May-Grunwald and Giemsa stains. The isolated cell population was significantly depleted by Mitomycin C administration, had a high RNA content, was proliferative, had a cytological profile that matched that of Piwi1+ stem cells, and was [~]10x enriched with Piwi1+ stem cells compared to whole cell suspension, all of which support the conclusion that the isolated population is indeed comprised of stem cells. This gene-agnostic FACS technique will serve future research into Hydractinia stem cell biology by enabling the use of isolated populations of live stem cells in transplantation, cell culture, and spheroid experimentation, and may serve as a reference for the development of new methods in other cnidarian species.

cell biology↗

bioRxiv2019

CRISPR-based platform for multimodal genetic screens in human iPSC-derived neurons

CRISPR/Cas9-based functional genomics have transformed our ability to elucidate mammalian cell biology. However, most previous CRISPR-based screens were conducted in cancer cell lines, rather than healthy, differentiated cells. Here, we describe a CRISPR interference (CRISPRi)-based platform for genetic screens in human neurons derived from induced pluripotent stem cells (iPSCs). We demonstrate robust and durable knockdown of endogenous genes in such neurons, and present results from three complementary genetic screens. First, a survival-based screen revealed neuron-specific essential genes and genes that improved neuronal survival upon knockdown. Second, a screen with a single-cell transcriptomic readout uncovered several examples of genes whose knockdown had strikingly cell-type specific consequences. Third, a longitudinal imaging screen detected distinct consequences of gene knockdown on neuronal morphology. Our results highlight the power of unbiased genetic screens in iPSC-derived differentiated cell types and provide a platform for systematic interrogation of normal and disease states of neurons.

cell biology↗

bioRxiv2026

Spatiotemporal Dynamics of Protein Recruitment During Cell Wound Repair

Injuries to individual cells happen frequently as a result of physiological and environmental stresses during their normal daily functions that can lead to a ruptured cell cortex (plasma membrane and underlying cortical cytoskeleton). The capacity of cells to rapidly repair general daily injuries, as well as ones resulting from trauma, infection, or diseases/cancer, is essential for their survival. While we know the general cell biological outline of the highly-conserved physiological events taking place during cell wound repair, our knowledge of the molecular mechanisms governing the repair process is still fairly limited, due in large part to the lack of information regarding the molecules, machineries, and pathways involved. Here, we performed a genetic screen of 1322 fluorescent-tagged proteins to identify cell wound repair components that are recruited upon laser wounding or whose expression is lost and/or altered upon laser wounding. We identified 129 proteins that are recruited to wounds during the cell repair process through high resolution spatio-temporal expression analyses of these gene fusions in conjunction with a fluorescent actin reporter. Strikingly, we find that many members of the Rab family GTPases are recruited to wounds where, in addition to their well-known roles in intracellular membrane trafficking, they are affecting actin cytoskeletal organization and dynamics during the repair process. These studies are allowing us to define the earliest acting proteins, as well as those required at specific steps in the repair process based on their recruitment patterns and the precise timing of their recruitment to wounds. Thus, our imaging-based screen is providing us with a global view of the repair processes, as well as a large number of genes/gene families that provide new entry points for examining specific steps in the cell wound repair process. Author SummaryCells in our bodies get injured every day from normal activity, environmental stress, infection, or disease. To survive, they must quickly repair these injuries and restore normal function. While some molecules have been identified as key players of cell wound repair, many of the molecules involved and their roles remain unknown. In this study, we identified new molecules that are involved in different steps of cell wound repair. Using laser-induced injury in the Drosophila model, we examined 1322 proteins and observed their spatial and temporal dynamics in a cell after injury. From the 1322 proteins examined, we identified 129 proteins recruited to distinct regions around the damage site during cell wound repair, suggesting roles in specific steps of the repair process. Interestingly, a subset of these proteins are Rab family GTPase members, highlighting new roles for these proteins in regulating actin dynamics. By identifying new candidate repair molecules, we provide a foundation for understanding how cells maintain their integrity and how repair processes may be influenced by factors such as wound size, infection, aging, and disease.

cell biology↗

bioRxiv2017

Revealing The Cell-Material Interface With Nanometer Resolution By FIB-SEM

The interface between biological cells and non-biological surfaces profoundly influences cellular activities, chronic tissue responses, and ultimately the success of medical implants. Materials in contact with cells can be plastics, metal, ceramics or other synthetic materials, and their surfaces vary widely in chemical compositions, stiffness, topography and levels of roughness. To understand the molecular mechanism of how cells and tissues respond to different materials, it is of critical importance to directly visualize the cell-material interface at the relevant length scale of nanometers. Conventional ultrastructural analysis by transmission electron microscopy (TEM) often requires substrate removal before microtome sectioning, which is not only challenging for most substrates but also can cause structural distortions of the interface. Here, we present a new method for in situ examination of the cell-to-material interface at any desired cellular location, based on focused-ion beam milling and scanning electron microscopy imaging (FIB-SEM). This method involves a thin-layer plastification procedure that preserves adherent cells as well as enhances the contrast of biological specimen. We demonstrate that this unique procedure allows the visualization of cell-to-material interface and intracellular structures with 10nm resolution, compatible with a variety of materials and surface topographies, and capable of volume and multi-directional imaging. We expect that this method will be very useful for studies of cell-to-material interactions and also suitable for in vivo studies such as examining osteoblast adhesion and new bone formation in response to titanium implants.

biophysics↗

bioRxiv2020

Discovery of membrane-permeating cyclic peptides via mRNA display

Small synthetic peptides capable of crossing biological membranes represent valuable tools in cell biology and drug delivery. While several cell-penetrating peptides (CPPs) of natural or synthetic origin have been reported, no peptide is currently known to cross both cytoplasmic and outer embryonic membranes. Here we describe a method to engineer membrane-permeating cyclic peptides (MPPs) with broad permeation activity by screening mRNA display libraries of cyclic peptides against embryos at different developmental stages. The proposed method was demonstrated by identifying peptides capable of permeating Drosophila melanogaster (fruit fly) embryos and mammalian cells. The selected peptide cyclo[Glut-MRKRHASRRE-K*] showed a strong permeation activity of embryos exposed to minimal permeabilization pretreatment, as well as human embryonic stem cells and a murine fibroblast cell line. Notably, in both embryos and mammalian cells, the cyclic peptide outperformed its linear counterpart and the control MPPs. Confocal microscopy and single cell flow cytometry analysis were utilized to assess the degree of permeation both qualitatively and quantitatively. These MPPs have potential application in studying and non-disruptively controlling intracellular or intraembryonic processes.

biochemistry↗

bioRxiv2020

Stable transfection in the protist Corallochytrium limacisporum allows identification of novel cellular features among unicellular relatives of animals

The evolutionary path from protists to multicellular animals remains a mystery. Recent work on the genomes of several unicellular relatives of animals has shaped our understanding of the genetic changes that may have occurred in this transition. However, the specific cellular modifications that took place to accommodate these changes remain unclear. Functional approaches are now needed to unravel how different cell biological features evolved. Recent work has already established genetic tools in three of the four unicellular lineages closely related to animals (choanoflagellates, filastereans, and ichthyosporeans). However, there are no genetic tools available for Corallochytrea, the lineage that seems to have the widest mix of fungal and metazoan features, as well as a complex life cycle. Here, we describe the development of stable transfection in the corallochytrean Corallochytrium limacisporum. Using a battery of cassettes to tag key cellular components, such as nucleus, plasma membrane, cytoplasm and actin filaments, we employ live imaging to discern previously unknown biological features of C. limacisporum. In particular, we identify two different paths for cell division--binary fission and coenocytic growth--that reveal a non-linear life cycle in C. limacisporum. Additionally, we found that C. limacisporum is binucleate for most of its life cycle, and that, contrary to what happens in most eukaryotes, nuclear division is decoupled from cell division. The establishment of these tools in C. limacisporum fills an important gap in the unicellular relatives of animals, opening up new avenues of research with broad taxon sampling to elucidate the specific cellular changes that occurred in the evolution of animals.

cell biology↗

bioRxiv2024

The 15-min (Sub)Cellular Proteome

Single-cell mass spectrometry (MS) opens a proteomic window onto the inner workings of cells. Here, we report the discovery characterization of the subcellular proteome of single, identified embryonic cells in record speed and molecular coverage. We integrated subcellular capillary microsampling, fast capillary electrophoresis (CE), high-efficiency nano-flow electrospray ionization, and orbitrap tandem MS. In proof-of-principle tests, we found shorter separation times to hinder proteome detection using DDA, but not DIA. Within a 15-min effective separation window, CE data-independent acquisition (DIA) was able to identify 1,161 proteins from single HeLa-cell-equivalent ([~]200 pg) proteome digests vs. 401 proteins by the reference data-dependent acquisition (DDA) on the same platform. The approach measured 1,242 proteins from subcellular niches in an identified cell in the live Xenopus laevis (frog) embryo, including many canonical components of organelles. CE-MS with DIA enables fast, sensitive, and deep profiling of the (sub)cellular proteome, expanding the bioanalytical toolbox of cell biology. Authorship ContributionsP.N. and B.S. designed the study. L.R.P. collected the X. laevis cell aspirates. B.S. prepared and measured the samples. B.S. and P.N. analyzed the data and interpreted the results. P.N. and B.S. wrote the manuscript. All the authors commented on the manuscript.

cell biology↗

bioRxiv2024

Laminin 111 triggers cell quiescence and long-term survival by inducing IQGAP1-mediated cytosolic scaffolding of ERK and BAD inactivation

In an adult human body, only a minority ([~]1%) of cells are dividing; all others are either quiescent, senescent or terminally differentiated. Cellular quiescence, also called G0, is a reversible non-proliferative state in which cells, such as adult stem cells, exist until stimuli trigger their re-entry into the cell cycle. Quiescent cells are known to reside within microenvironment niches of specific extracellular matrix (ECM) composition, but the molecular mechanisms that control their entry and maintenance into G0 and their long-term survival are poorly understood. Here, using a reproducible and homogenous in vitro model of quiescence, ex vivo tissue histology, phosphoproteomics, and molecular cell biological assays, we revealed that Laminin 111 was sufficient to trigger i) reversible cell cycle exit into G0; ii) sustained and elevated MAPK/ERK signaling; and iii) long-term survival. We found that ERK was activated through the Rap1-BRAF-MEK arm underneath Laminin-binding Integrin 3{beta}1. Activated pERK was scaffolded into the cytoplasm by IQGAP1, thereby blocking its translocation into the nucleus and the activation of proliferative transcription factors. Instead, cytoplasmic pERK inhibited pro-apoptotic protein BAD, which mediated the survival of quiescent cells even in absence of mitogen stimuli. Importantly, we confirmed that pERK was elevated and retained in the cytoplasm of Lgr5+ stem cells when they were located within Laminin 1-positive niches in porcine intestine. These findings uncovered a molecular mechanism that may explain how quiescent cell pools, such as dormant adult stem cells, can survive many years despite low mitogen stimuli and be resistant to apoptotic challenges, including chemotherapy. HIGHLIGHTSO_LILaminin 111 is sufficient to induce cellular quiescence (G0) and long-term survival. C_LIO_LILaminin 111 triggers the sustained and elevated activation of ERK during G0. C_LIO_LIERK is activated not by growth factor receptors but through the Rap1-BRAF-MEK arm underneath Laminin-binding Integrin 3{beta}1. C_LIO_LIActive, phosphorylated ERK (pERK) is scaffolded by IQGAP1, which prevents it from translocating into the nucleus and activating proliferative transcription factors. C_LIO_LIInstead, cytoplasmic pERK mediates the phosphorylation, and thus inhibition, of BAD, thereby raising the threshold at which G0 cells enter apoptosis. C_LI

cell biology↗

bioRxiv2025

WS6 enables scalable ex vivo expansion and gene editing of epithelial basal stem cells

Modeling human epithelial diseases and developing cell-based therapies require robust methods to expand and manipulate epithelial stem and progenitor cells in vitro. Basal stem/progenitor cells from stratified epithelia can be expanded in 3T3-J2 fibroblast feeder cell co-culture systems, and the addition of the ROCK inhibitor Y-27632 enhances proliferation and culture longevity, a phenomenon described as conditional reprogramming. Here, we present a method incorporating the small molecule WS6 to further improve the proliferation and lifespan of cultured epithelial cells from multiple tissues, including airway, skin, and thymus. Cells maintained in this medium ( EpMED; FAD+Y+WS6) retain basal stem/progenitor cell identity and function, including the capacity to differentiate. We demonstrate their capacity to engraft in vivo in a tracheal transplantation model. In a second application, we generate clonal CRISPR-Cas9 genome edited nasal cultures, introducing targeted knockouts of DNAH5 or DNAI2 to create primary ciliary dyskinesia disease models. We anticipate that our method will have broad applications in epithelial cell biology, disease modeling, and regenerative medicine, while reducing reliance on immortalized or cancer cell lines and animal experimentation.

cell biology↗

bioRxiv2017

Cytosolic delivery of large supramolecular proteincomplexes arranged on DNA nanopegboards

A generic methodology for cytosolic delivery of large supramolecular multiprotein complexes into living cells is described that takes advantage of the highly-controllable bottom-up fabrication of protein-decorated DNA nanostructures and the microfluidic \"cell squeezing\" technique. Therein, cells are deformed upon passage through a narrow constriction leading to formation of transient holes in the cell membrane that enable the diffusion of the protein-DNA nanostructures from the surrounding buffer into the cytosol. A diverse set of multiprotein complexes was assembled on DNA origami nanostructures using streptavidin and the sensitive glucose sensor protein FLIP as model systems. We demonstrate that our approach allows for the direct cytosolic delivery of these multifunctional protein complexes into the cytosol of HeLa cells. We also demonstrate that targeting groups can be incorporated into the protein-DNA nanoassemblies to enable their intracellular targeting to cytosolic compartments, such as the cytoskeleton or nucleus. We believe that this methodology will open up novel strategies for research in fundamental cell biology, such as the reverseengineering of the supramolecular machinery involved in gene regulation, cell signalling, or cell division. Furthermore, direct applications in immunotherapy can be foreseen.

cell biology↗

bioRxiv2021

A cellular and molecular atlas reveals the basis of chytrid development.

The chytrids (phylum Chytridiomycota) are a major early-diverging fungal lineage of ecological and evolutionary importance. Despite their importance, many fundamental aspects of chytrid developmental and cell biology remain poorly understood. To address these knowledge gaps, we combined quantitative volume electron microscopy and comparative transcriptome profiling to create an atlas of the cellular and molecular basis of the chytrid life cycle, using the model chytrid Rhizoclosmatium globosum. From our developmental atlas, we show that zoospores exhibit a specialised biological repertoire dominated by inactive ribosome aggregates, and that lipid processing is complex and dynamic throughout the cell cycle. We demonstrate that the chytrid apophysis is a distinct subcellular structure characterised by high intracellular trafficking, providing evidence for division of labour in the chytrid cell plan, and show that zoosporogenesis includes animal like amoeboid cell morphologies resulting from endocytotic cargo transport from the interstitial maternal cytoplasm. Taken together, our results reveal insights into chytrid developmental biology and provide a basis for future investigations into early-diverging fungal cell biology.

developmental biology↗

bioRxiv2022

Found in translation: Microproteins are a new class of potential host cell impurity in mAb drug products

Chinese hamster ovary (CHO) cells are used to produce almost 90% of therapeutic monoclonal antibodies (mAbs). The annotation of non-canonical translation events in these cellular factories remains incomplete, limiting not only our ability to study CHO cell biology but also detect host cell protein (HCP) contaminants in the final mAb drug product. We utilised ribosome footprint profiling (Ribo-seq) to identify novel open reading frames (ORFs) including N-terminal extensions and thousands of short ORFs (sORFs) predicted to encode microproteins. Mass spectrometry-based HCP analysis of four commercial mAb drug products using the extended protein sequence database revealed the presence of microprotein impurities for the first time. We also show that microprotein abundance varies with growth phase and can be affected by the cell culture environment. In addition, our work provides a vital resource to facilitate future studies of non-canonical translation as well as the regulation of protein synthesis in CHO cell lines.

genomics↗

bioRxiv2022

The Structure-Selective Endonucleases GEN1 and MUS81 are Functionally Complementary in Safeguarding the Genome of Proliferating B Lymphocytes

During the development of humoral immunity, activated B lymphocytes undergo vigorous proliferative, transcriptional, metabolic, and DNA remodeling activities; hence, their genomes are constantly exposed to an onslaught of genotoxic agents and processes. Recombination-dependent DNA transactions that preserve the integrity of the genome and of the DNA replication process generates Holliday junctions that must be eliminated for the accurate segregation of sister chromatids and faithful propagation of genomic material. To investigate the role of two Holliday junction resolvases, GEN1 and MUS81, in B cell biology, we established B-cell conditional knockout mouse models and found that targeted deletion of GEN1 and MUS81 in early B cell precursors halts their development and maturation while selective loss of the resolvases in mature B cells inhibits the generation of robust germinal centers. Upon activation, these double-null mature B lymphocytes fail to proliferate and survive while exhibiting transcriptional signatures of p53 signaling, apoptosis, and type I interferon response. Metaphase spreads of these resolvase-deficient cells showed severe and diverse chromosomal abnormalities, including a preponderance of chromosome breaks, consistent with a defect in resolving DNA recombination intermediates. These observations underscore the essential roles of GEN1 and MUS81 in safeguarding the genome to ensure the proper development and maintenance of B lymphocytes.

immunology↗

bioRxiv2023

Plasmid-free cheater cells commonly evolve during laboratory growth

It has been nearly a century since the isolation and use of penicillin, heralding the discovery of a wide range of different antibiotics. In addition to clinical applications, such antibiotics have been essential laboratory tools, allowing for selection and maintenance of laboratory plasmids that encode cognate resistance genes. However, antibiotic resistance mechanisms can additionally function as public goods. For example, secretion of beta-lactamase from resistant cells, and subsequent degradation of nearby penicillin and related antibiotics, allows neighboring plasmid-free susceptible bacteria to survive antibiotic treatment. How such cooperative mechanisms impact selection of plasmids during experiments in laboratory conditions is poorly understood. Here, we show that the use of plasmid-encoded beta-lactamases leads to significant curing of plasmids in surface grown bacteria. Furthermore, such curing was also evident for aminoglycoside phosphotransferase and tetracycline antiporter resistance mechanisms. Alternatively, antibiotic selection in liquid growth led to more robust plasmid maintenance, although plasmid loss still occurred. The net outcome of such plasmid loss is the generation of a heterogenous population of plasmid-containing and plasmid-free cells, leading to experimental confounds that are not widely appreciated. ImportancePlasmids are routinely used in microbiology as readouts of cell biology or tools to manipulate cell function. Central to these studies is the assumption that all cells in an experiment contain the plasmid. Plasmid maintenance in a host cell typically depends on a plasmid-encoded antibiotic resistance marker, which provides a selective advantage when the plasmid containing cell is grown in the presence of antibiotic. Here we find that growth of plasmid-containing bacteria during laboratory conditions in the presence of three distinct antibiotic families leads to the evolution of a significant number of plasmid-free cells, which rely on the resistance mechanisms of the plasmid-containing cells for viability. This process generates a heterogenous population of plasmid-free and plasmid-containing bacteria, an outcome which could confound further experimentation.

microbiology↗

bioRxiv2024

Leveraging Crosslinker Diffusion to Template Stiffness Gradients in Alginate Hydrogels

Mechanobiology drives many important cell biological behaviors such as stem cell differentiation, cancer drug resistance and cell migration up stiffness gradients, a process called durotaxis. The development of 3D hydrogel systems with tunable 2D mechanical gradient patterns affords the ability to study these mechanosensitive cell behaviors to understand cancer invasion or enhance wound healing through directed migration. In this paper, we developed an approach to spatially imprint within alginate hydrogels, gradients in mechanical properties that can be used to probe mechanobiology. Stencils were easily designed and fabricated using a common craft cutter to control the presentation of a calcium crosslinking solution to alginate gels. Different stencil shapes result in different gradients in opacity that can be imprinted into both thick and thin alginate gels of arbitrary 2D shape. The steepness of the opacity gradient as well as the maximum opacity can be controlled based on reproducible crosslinking kinetics regulated through calcium concentration and gradient developing time. Calcium crosslinking results in both opacity changes as well as increases in elastic modulus in the bulk hydrogel. Opacity correlates with elastic modulus, allowing it to be used as a proxy for local elastic modulus. Functionalized alginate gels with collagen and imprinting stiffness gradients within them resulted in cell invasion that was spatially dependent, where stiffer regions facilitated deeper invasion of breast cancer cells. Consequently, this stenciling approach represents a facile way to control stiffness gradients in alginate gels in order to study mechanosensitive cellular behavior. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=45 SRC="FIGDIR/small/599742v2_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@1182404org.highwire.dtl.DTLVardef@1e4bfb1org.highwire.dtl.DTLVardef@e52778org.highwire.dtl.DTLVardef@8237bd_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2026

Cell-level random splits leak group-owned answers in single-cell benchmarks

Machine learning models in single-cell biology increasingly forecast differentiation, reprogramming and therapeutic response from early transcriptomic profiles. Testing whether a model has learned real biology requires held-out cells. Single-cell data, however, are grouped: cells from the same clone, patient or batch share the same label. A random split therefore places relatives of each test cell, carrying its label, in the training set, and a model can score well by memorizing a relative instead of learning a transferable rule. Grouped validation removes this leakage but leaves far fewer independent units behind each error bar. Here we show how to estimate this leakage before training any model, from two properties of the data: exposure, the fraction of test cells with relatives in training, and retrievability, how often a nearest-neighbor search returns such a relative rather than an unrelated cell. Across lineage-barcoded and patient data, exposure determines whether a random split opens a leakage channel, and retrievability determines how much it can inflate the score. The inflation is negligible where cell state has decoupled from ancestry, much larger where clonal sisters remain close in expression space, and in a patient cohort large enough to overturn a clinical conclusion. We also provide eakcheck, which computes both properties in seconds, before the outcome model is fitted.

bioinformatics↗

bioRxiv2019

Fungal Feature Tracker (FFT): A tool for quantitatively characterizing the morphology and growth of filamentous fungi

Filamentous fungi are ubiquitous in nature and serve as important biological models in various scientific fields including genetics, cell biology, ecology, evolution, and chemistry. A significant obstacle in studying filamentous fungi is the lack of tools for characterizing their growth and morphology in an efficient and quantitative manner. Consequently, assessments of the growth of filamentous fungi are often subjective and imprecise. In order to remedy this problem, we developed Fungal Feature Tracker (FFT), a user-friendly software comprised of different image analysis tools to automatically quantify different fungal characteristics, such as spore number, spore morphology, and measurements of total length, number of hyphal tips and the area covered by the mycelium. In addition, FFT can recognize and quantify specialized structures such as the traps generated by nematode-trapping fungi, which could be tuned to quantify other distinctive fungal structures in different fungi. We present a detailed characterization and comparison of a few fungal species as a case study to demonstrate the capabilities and potential of our software. Using FFT, we were able to quantify various features at strain and species level, such as mycelial growth over time and the length and width of spores, which would be difficult to track using classical approaches. In summary, FFT is a powerful tool that enables quantitative measurements of fungal features and growth, allowing objective and precise characterization of fungal phenotypes.\n\nAuthor SummaryOne of the main obstacles to study filamentous fungi is the lack of tools for characterizing fungal phenotypes in an efficient and quantitative manner. Assessment of cell growth and numbers rely on tedious manual techniques that often result in subjective and imprecise measurements. In response to those limitations, we developed Fungal Feature Tracker (FFT), a user-friendly software that allows researchers to characterize different phenotypic features of filamentous fungi such as sporulation, spore morphology and mycelial growth. In addition, FFT can recognize and quantify other fungal structures including the fungal traps developed by nematode-trapping fungi. In order to show the capabilities and potential of our software, we conducted a detailed characterization and comparison of different fungal species. Our comparison relies on a series of experimental set-ups using standard and easily accessible equipment to ensure reproducibility in other laboratories. In summary, FFT is an easy to use and powerful tool that can quantitatively characterize fungal morphology, cell number and quantitatively measures the filamentous growth, which will allow advance our understanding of the growth and biology of filamentous fungi.

cell biology↗